Advertisement

Identification of Arabidopsis Mutants with Altered Freezing Tolerance

  • Carlos Perea-Resa
  • Rafael Catalá
  • Julio SalinasEmail author
Protocol
  • 13 Downloads
Part of the Methods in Molecular Biology book series (MIMB, volume 2156)

Abstract

Low temperature is an important determinant in the configuration of natural plant communities and defines the range of distribution and growth of important crops. Some plants, including Arabidopsis thaliana, have evolved sophisticated adaptive mechanisms to tolerate freezing temperatures. Central to this adaptation is the process of cold acclimation. By means of this process, many plants from temperate regions are able to develop or increase their freezing tolerance in response to low, nonfreezing temperatures. The identification and characterization of factors involved in freezing tolerance is crucial to understand the molecular mechanisms underlying the cold acclimation response and has a potential interest to improve crop tolerance to freezing temperatures. Many genes implicated in cold acclimation have been identified in numerous plant species by using molecular approaches followed by reverse genetic analysis. Remarkably, however, direct genetic analyses have not been conveniently exploited in their capacity for identifying genes with pivotal roles in that adaptive response. In this chapter, we describe a protocol for evaluating the freezing tolerance of both nonacclimated and cold acclimated Arabidopsis plants. This protocol allows for the accurate and simple screening of mutant collections for the identification of novel factors involved in freezing tolerance and cold acclimation.

Key words

Freezing temperature Freezing tolerance Cold acclimation Arabidopsis thaliana 

References

  1. 1.
    Levitt J (1980) Responses of plants to environmental stresses: chilling, freezing and high temperatures stresses, 2nd edn. Academic, New YorkGoogle Scholar
  2. 2.
    Steponkus PL, Uemura M, Joseph RA et al (1998) Mode of action of the COR15a gene on the freezing tolerance of Arabidopsis thaliana. Proc Natl Acad Sci U S A 95:14570–14575CrossRefGoogle Scholar
  3. 3.
    Guy CL (1990) Cold acclimation and freezing stress tolerance: role of protein metabolism. Annu Rev Plant Physiol Plant Mol Biol 41:187–223CrossRefGoogle Scholar
  4. 4.
    Lightner J, Caspar T (1998) Seed mutagenesis of Arabidopsis. In: Martínez-Zapater JM, Salinas J (eds) Methods in molecular biology, vol 82. Humana, Totowa, NJ, pp 91–103Google Scholar
  5. 5.
    Kim YS, Schumaker KS, Zhu JK (2006) EMS mutagenesis of Arabidopsis. In: Salinas J, Sanchez-Serrano JJ (eds) Methods in molecular biology, vol 323. Humana, Totowa, NJ, pp 101–103Google Scholar
  6. 6.
    Weigel D, Glazebrook J (2006) EMS mutagenesis of Arabidopsis seed. CSH Protoc 28.  https://doi.org/10.1101/pdb.prot4621
  7. 7.
    Warren G, McKown R, Marin AL et al (1996) Isolation of mutations affecting the development of freezing tolerance in Arabidopsis thaliana (L.) Heynh. Plant Physiol 111:1011–1019CrossRefGoogle Scholar
  8. 8.
    McKown R, Kuroki G, Warren G (1996) Cold responses of Arabidopsis mutants impaired in freezing tolerance. J Exp Bot 47:1919–1925CrossRefGoogle Scholar
  9. 9.
    Xin Z, Browse J (1998) eskimo1 mutants of Arabidopsis are constitutively freezing-tolerant. Proc Natl Acad Sci U S A 95:7799–7804CrossRefGoogle Scholar
  10. 10.
    Llorente F, Oliveros JC, Martínez-Zapater JM et al (2000) A freezing-sensitive mutant of Arabidopsis, frs1, is a new aba3 allele. Planta 211:648–655CrossRefGoogle Scholar
  11. 11.
    Koornneef M, Dellaert LWM, van der Veen JH (1982) EMS- and radiation-induced mutation frequencies at individual loci in Arabidopsis thaliana (L.) Heynh. Mutat Res 93:109–123CrossRefGoogle Scholar
  12. 12.
    Alonso JM, Stepanova AN (2003) T-DNA mutagenesis in Arabidopsis. In: Grotewold E (ed) Methods in molecular biology, vol 236. Humana, Totowa, NJ, pp 177–188Google Scholar
  13. 13.
    Jander G (2006) Gene identification and cloning by molecular marker mapping. In: Salinas J, Sanchez-Serrano JJ (eds) Methods in molecular biology, vol 323. Humana, Totowa, NJ, pp 115–126Google Scholar
  14. 14.
    Austin RS, Vidaurre D, Stamatiou G et al (2011) Next-generation mapping of Arabidopsis genes. Plant J 67:715–725CrossRefGoogle Scholar
  15. 15.
    Uchida N, Sakamoto T, Kurata T et al (2011) Identification of EMS-induced causal mutations in a non-reference Arabidopsis thaliana accession by whole genome sequencing. Plant Cell Physiol 52:716–722CrossRefGoogle Scholar
  16. 16.
    Wachsman G, Modliszewski JL, Valdes M, Benfey PN (2017) A SIMPLE pipeline for mapping point mutations. Plant Physiol 174:1307–1313CrossRefGoogle Scholar
  17. 17.
    O’Malley RC, Alonso JM, Kim CJ et al (2007) An adapter ligation-mediated PCR method for high-throughput mapping of T-DNA inserts in the Arabidopsis genome. Nat Protoc 2:2910–2917CrossRefGoogle Scholar
  18. 18.
    McCourt P, Keith K (1998) Sterile techniques in Arabidopsis, Methods in molecular biology, vol 82. Humana, Totowa, NJ, pp 13–17Google Scholar
  19. 19.
    Mikkelsen MD, Thomashow MF (2009) A role for circadian evening elements in cold-regulated gene expression in Arabidopsis. Plant J 60:328–339CrossRefGoogle Scholar
  20. 20.
    Dong MA, Farré EM, Thomashow MF (2011) Circadian clock-associated 1 and late elongated hypocotyl regulate expression of the C-repeat binding factor (CBF) pathway in Arabidopsis. Proc Natl Acad Sci U S A 108:7241–7246CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Carlos Perea-Resa
    • 1
  • Rafael Catalá
    • 1
  • Julio Salinas
    • 1
    Email author
  1. 1.Departamento de Biotecnología Microbiana y de PlantasCentro de Investigaciones Biológicas Margarita Salas (CIB-CSIC)MadridSpain

Personalised recommendations